title: A one-step reverse transcription loop-mediated isothermal amplification for detection and discrimination of infectious bursal disease virus.
abstract: BACKGROUND
  Infectious bursal disease (IBD) is a highly contagious immunosuppressive disease in young chickens caused by infectious bursal disease virus (IBDV). It causes huge economic losses to the poultry industry. The objective of this study is to develop a loop-mediated isothermal amplification (LAMP) method for the detection and discrimination of IBDV.
  
  
  RESULTS
  In this study, we applied reverse transcription loop-mediated isothermal amplification (RT-LAMP) to detect IBDV in one simple step and further identified the very virulent strain from non-vvIBDVs with a simply post-amplification restriction enzyme analysis. Based on sequence analysis, a set of two inner, two outer and two loop primers were designed to target the VP5 gene and they showed great specificity with no cross reaction to the other common avian pathogens. The detection limit determined by both color change inspection and agarose gel electrophoresis was 28 copies viral RNA, which was almost as sensitive as a real-time RT-PCR previous developed in our laboratory. We also identified a unique Tfi I restriction site located exclusively in non-vvIBDVs, so very virulent strain could be distinguished from current vaccine strains. By screening a panel of clinical specimens, results showed that this method is high feasible in clinical settings, and it obtained results 100% correlated with real-time RT-PCR.
  
  
  CONCLUSIONS
  RT-LAMP is a rapid, simple and sensitive assay. In combination with the Tfi I restriction analysis, this method holds great promises not only in laboratory detection and discrimination of IBDV but also in large scale field and clinical studies.
authors.affiliation: Division of Avian Infectious Diseases, State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin, Heilongjiang 150001, China.
authors.forename: Yongqiang
authors.initials: Y
authors.lastname: Wang
authors.forename: Zhonghui
authors.initials: Z
authors.lastname: Kang
authors.forename: Honglei
authors.initials: H
authors.lastname: Gao
authors.forename: Yulong
authors.initials: Y
authors.lastname: Gao
authors.forename: Liting
authors.initials: L
authors.lastname: Qin
authors.forename: Huan
authors.initials: H
authors.lastname: Lin
authors.forename: Fei
authors.initials: F
authors.lastname: Yu
authors.forename: Xiaole
authors.initials: X
authors.lastname: Qi
authors.forename: Xiaomei
authors.initials: X
authors.lastname: Wang
mesh_terms: D000818:Animals; D018175:Birnaviridae Infections / Q000175:diagnosis / Q000662:veterinary* / Q000821:virology; D002642:Chick Embryo; D002645:Chickens; D017931:DNA Primers / Q000235:genetics; D007243:Infectious bursal disease virus / Q000235:genetics / Q000302:isolation & purification*; D008969:Molecular Sequence Data; D011201:Poultry Diseases / Q000175:diagnosis* / Q000821:virology; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D014764:Viral Proteins / Q000235:genetics
chemical_list: D017931:DNA Primers; D014764:Viral Proteins